Medel Manuel L. Zulueta

Synthesis of 3-O-sulfonated heparan sulfate octasaccharides that inhibit the herpes simplex virus type 1 host–cell interaction

Cell surface carbohydrates play significant roles in a number of biologically important processes. Heparan sulfate, for instance, is a ubiquitously distributed polysulfated polysaccharide that is involved, among other things, in the initial step of herpes simplex virus type 1 (HSV-1) infection. The virus interacts with cell-surface heparan sulfate to facilitate host-cell attachment and entry. 3-O-Sulfonated heparan sulfate has been found to function as an HSV-1 entry receptor. Achieving a complete understanding of these interactions requires the chemical synthesis of such oligosaccharides, but this remains challenging. Here, we present a convenient approach for the synthesis of two irregular 3-O-sulfonated heparan sulfate octasaccharides, making use of a key disaccharide intermediate to acquire different building blocks for the oligosaccharide chain assembly. Despite substantial structural differences, the prepared 3-O-sulfonated sugars blocked viral infection in a dosage-dependent manner with remarkable similarity to one another.

α-Glycosylation by d-Glucosamine-Derived Donors: Synthesis of Heparosan and Heparin Analogues That Interact with Mycobacterial Heparin-Binding Hemagglutinin

Numerous biomolecules possess α-D-glucosamine as structural component. However, chemical glycosylations aimed at this backbone are usually not easily attained without generating the unwanted β-isomer. We report herein a versatile approach in affording full α-stereoselectivity built upon a carefully selected set of orthogonal protecting groups on a D-glucosaminyl donor. The excellent stereoselectivity provided by the protecting group combination was found independent of leaving groups and activators. With the trichloroacetimidate as the optimum donor leaving group, core skeletons of glycosylphosphatidyl inositol anchors, heparosan, heparan sulfate, and heparin were efficiently assembled. The orthogonal protecting groups were successfully manipulated to further carry out the total syntheses of heparosan tri- and pentasaccharides and heparin di-, tetra-, hexa-, and octasaccharide analogues. Using the heparin analogues, heparin-binding hemagglutinin, a virulence factor of Mycobacterium tuberculosis, was found to bind at least six sugar units with the interaction notably being entropically driven.

Divergent synthesis of 48 heparan sulfate-based disaccharides and probing the specific sugar-fibroblast growth factor-1 interaction.

Several biological processes involve glycans, yet understanding their ligand specificities is impeded by their inherent diversity and difficult acquisition. Generating broad synthetic sugar libraries for bioevaluations is a powerful tool in unraveling glycan structural information. In the case of the widely distributed heparan sulfate (HS), however, the 48 theoretical possibilities for its repeating disaccharide call for synthetic approaches that should minimize the effort in an undoubtedly huge undertaking. Here we employed a divergent strategy to afford all 48 HS-based disaccharides from just two orthogonally protected disaccharide precursors. Different combinations and sequence of transformation steps were applied with many downstream intermediates leading up to multiple target products. With the full disaccharide library in hand, affinity screening with fibroblast growth factor-1 (FGF-1) revealed that four of the synthetic sugars bind to FGF-1. The molecular details of the interaction were further clarified through X-ray analysis of the sugar-protein cocrystals. The capability of comprehensive sugar libraries in providing key insights in glycan-ligand interaction is, thus, highlighted.

Acyl and Silyl Group Effects in Reactivity-Based One-Pot Glycosylation: Synthesis of Embryonic Stem Cell Surface Carbohydrates Lc4 and IV2Fuc-Lc4

Relative reactivity evaluations showed the graded arming of toluenyl thioglucosides by variously positioned silyl groups but not by their acyl counterparts. These findings were applied in reactivity-based one-pot assembly of linker-attached Lc(4) and IV(2)Fuc-Lc(4), which are components of human embryonic stem cell surface. The sugar-galectin-1 binding was also examined.

Regioselective One-Pot Protection of D-Glucosamine

A highly regioselective one-pot transformation of 2-azido-2-deoxy-1,3,4,6-tetra-O-trimethylsilyl-d-glucopyranose via sequential additions of various reagents was systematically studied, yielding the fully protected derivatives and the 1-, 3-, 4-, as well as 6-alcohols, respectively.

Synthetic heparin and heparan sulfate oligosaccharides and their protein interactions.

Heparin and heparan sulfate bind a host of basic proteins that take advantage of the sugars dense structural information. The significance of these interactions in various aspects of development, physiology, and disease stimulated keen interest in evaluating structure-activity relationships. The well-defined heparin and heparan sulfate oligosaccharides needed for these studies can be mainly accessed by chemical synthesis and, more recently by chemoenzymatic means. The various synthetic strategies available to chemical synthesis have recently enabled the acquisition of several regular and irregular sequences, including a number of dodecasaccharides, through improved coupling methods and judicial protecting group manipulations. Controlled chain elongation and critical application of modification enzymes allowed the generation of well-defined constructs via chemoenzymatic synthesis. Investigations of various protein interactions with the synthetic constructs delivered valuable information that could aid future drug development endeavors.

One-Pot Strategies for the Synthesis of the Tetrasaccharide Linkage Region of Proteoglycans

A linker-attached tetrasaccharide corresponding to the linkage region of proteoglycans was synthesized via one-pot procedures from the silylated monosaccharide derivatives. Regioselective one-pot protection protocols were applied in generating the requisite monosaccharide building blocks whereas stereoselective one-pot glycosylation approaches were utilized to assemble the tetrasaccharide skeleton.

Structural basis for oligomerization and glycosaminoglycan binding of CCL5 and CCL3.

Significance Oligomerization and glycosaminoglycan (GAG) binding are key regulatory steps for many extracellular ligands. Our analyses provide a structural basis of CC chemokine ligand 5 (CCL5) and CCL3 oligomerization and explain how oligomerization affects the interaction of these chemokines with GAG and their functions. Our GAG-bound chemokine structures reveal how CCL5 and CCL3 oligomerization creates distinctive GAG-binding grooves to enhance GAG binding via avidity for regulating chemokine functions. Furthermore, our CCL5 structure may explain how CXCL4, a CXC chemokine, heterooligomerizes with CCL5 to modulate chemokine-mediated activities. Together, these data provide new structural insights into how oligomerization and GAG binding are coupled to regulate functions of CC chemokines and offer novel pharmacophores for the design of therapeutics for treating chemokine-mediated human diseases. CC chemokine ligand 5 (CCL5) and CCL3 are critical for immune surveillance and inflammation. Consequently, they are linked to the pathogenesis of many inflammatory conditions and are therapeutic targets. Oligomerization and glycosaminoglycan (GAG) binding of CCL5 and CCL3 are vital for the functions of these chemokines. Our structural and biophysical analyses of human CCL5 reveal that CCL5 oligomerization is a polymerization process in which CCL5 forms rod-shaped, double-helical oligomers. This CCL5 structure explains mutational data and offers a unified mechanism for CCL3, CCL4, and CCL5 assembly into high-molecular-weight, polydisperse oligomers. A conserved, positively charged BBXB motif is key for the binding of CC chemokines to GAG. However, this motif is partially buried when CCL3, CCL4, and CCL5 are oligomerized; thus, the mechanism by which GAG binds these chemokine oligomers has been elusive. Our structures of GAG-bound CCL5 and CCL3 oligomers reveal that these chemokine oligomers have distinct GAG-binding mechanisms. The CCL5 oligomer uses another positively charged and fully exposed motif, KKWVR, in GAG binding. However, residues from two partially buried BBXB motifs along with other residues combine to form a GAG-binding groove in the CCL3 oligomer. The N termini of CC chemokines are shown to be involved in receptor binding and oligomerization. We also report an alternative CCL3 oligomer structure that reveals how conformational changes in CCL3 N termini profoundly alter its surface properties and dimer–dimer interactions to affect GAG binding and oligomerization. Such complexity in oligomerization and GAG binding enables intricate, physiologically relevant regulation of CC chemokine functions.

Synthesis of L-hexoses and their related biomolecules

Carbohydrates either conjugated or as free entities are major players in numerous biological processes. The desire to comprehend the nature of their functions and further develop therapeutic and diagnostic applications has fuelled the recent upsurge in the glycoscience field. Mainly accessed through chemical synthesis, homogeneous and well-defined sugar constructs are on high demand for structure-activity evaluation. Although the d-sugars, particularly the d-hexoses, have dominated the carbohydrate landscape, L-hexoses also attracted attention because they are known components of important polysaccharides, antibiotics, and other natural products. Nonetheless, the L-hexose-based materials needed for making building blocks for sugar assemblies are rare and are usually expensive if commercially available. Thus, intense efforts were focused on the development of innovative and reliable methods for the acquisition of L-hexoses and their derivatives. This review outlines several efficient and cost-effective routes for the chemical syntheses of L-hexoses, particularly focusing on approaches that utilize commercially abundant sugars as starting materials. A sampling of the applications of the generated L-hexoses in preparing biologically relevant compounds is also provided.

Regioselective and stereoselective benzylidene installation and one-pot protection of D-mannose.

Oligosaccharide syntheses are an important source of well-defined sugar constructs particularly needed for the evaluation of structure-activity relationships. The chemical assembly of oligosaccharides requires several building blocks, that is, glycosyl donors and acceptors, which are prepared in multistep processes and in a generally tedious and time-consuming manner. Having developed one-pot procedures meant to minimise the effort in sugar building block preparation, we tackled herein the one-pot preparation of fully protected and 2-, 3-, 4-, and 6-alcohol derivatives of d-mannose, a widely distributed monosaccharide. As a consequence of the hydroxyl group pattern of D-mannose, regioselective and stereoselective benzylidenations were developed and later seamlessly utilised as the first transformation in the one-pot procedure.